The long-term goal of this proposal is to evaluate a physiological and therapeutic role for the renal angiotensin II (Ang II) AT2 receptors in improving renal/cardiovascular function and treating hypertension associated with obesity/diabetes. Of the two Ang II receptors, AT1 is known to mediate most of the Ang II actions such as antinatriuresis and increased blood pressure (BP). The AT2 receptor is suggested to function as an antagonist of AT1 receptors; however, low expression of the AT2 receptor has hindered development of a clear understanding of the physiological and therapeutic roles of this receptor. During the previous funding period, we found that hyperglycemia, associated with either insulin resistance or insulin insufficiency, caused an increase in AT2 receptor expression in the renal proximal tubule. Selective activation of the AT2 receptors stimulated a nitric oxide/cGMP pathway, thus inhibiting proximal tubule Na,K-ATPase (NKA) activity and increasing urinary Na excretion in obese Zucker rats, a model of insulin resistance and hyperglycemia. Treatment of obese rats with an AT2 receptor antagonist for 2 weeks raised BP in these animals. Three hypotheses were developed from these observations. First, hyperglycemia increases AT2 receptor expression by transcriptional activation. Second, selective AT2 receptor activation directly inhibits NKA via a NO/cGMP pathway and also, antagonizes the AT1 receptor-mediated NKA stimulation in proximal tubules by cGMP- mediated inhibition of phosphodiesterase-3 (PDE-3). Third, long-term selective AT2 receptor activation increases urinary Na excretion and lowers BP in obese Zucker/diabetic rats. These hypotheses will be tested by 3 specific aims utilizing cell culture and whole animal models. Aim 1 will study the role of p38 MAP kinase and transcription factors IRF-1 and C/EBP2 in increasing AT2 receptor expression by high glucose concentrations in HK-2 cells; renal AT2 receptor expression/function in obese rats with euglycemia. Aim 2 will study the role of cGMP-mediated PDE-3 inhibition, as a novel mechanism responsible for AT2 receptor antagonism of AT1 receptor-stimulated NKA, in isolated proximal tubules of obese Zucker rats. In aim 3, we will study the effect of long-term treatment with an AT2 agonist on renal function and BP in control in kidney- specific AT2 knock-down obese/lean Zucker rats. Following treatments, proximal tubule NKA activity and various components of the renin angiotensin system will be determined. Biochemical, immunological, molecular biology, fluorescence imaging and whole animal physiological methodologies will be employed to accomplish the proposed studies. These studies will elucidate the mechanisms regulating AT2 receptor in diabetes. Moreover, they will advance our understanding of the physiological role of the AT2 receptor in the long-term regulation of renal function and blood pressure control. Finally, the studies may clarify the potential of the AT2 receptor as an important therapeutic target providing a strong basis to develop selective non-peptide AT2 agonists.